Microfluidic devices and methods

Abstract

Contemplated microfluidic devices and methods are drawn to protein arrays in which distinct and detergent-containing antigen preparations are deposited onto an optical contrast layer in a non-specific and non-covalent manner. Detection of binding a is carried out using a dye that precipitates or agglomerates to so form a visually detectable signal at a dynamic range of at least three orders of magnitude.

What is claimed is: 
     
       1. A microfluidic device, comprising:
 an enclosed reaction volume between 1 μl and 50 μl, formed at least in part by a non-permeable glass or transparent polymer carrier material, and wherein the reaction volume is configured to allow retention of a solution comprising an antibody; 
 a porous optical contrast layer having a thickness of between 10-100 microns and coupled to a surface of the glass or transparent polymer carrier material, wherein the optical contrast layer is disposed within the reaction volume and opposite to a cavity layer within the reaction volume, and wherein the cavities in the cavity layer are circular cavities arranged at regular intervals in x- and y-coordinate; 
 wherein a plurality of distinct antigens are non-covalently and non-specifically bound to the optical contrast layer as antigen spots in respective predetermined positions, and wherein the optical contrast layer comprises a nontransparent material; 
 wherein the cavity layer is configured to provide a plurality of cavities having a radius of 400 μm or less and configured to allow trapping of air in the plurality of cavities, and wherein the plurality of cavities are (a) sized and positioned to allow acoustic air bubble resonance mixing of a fluid that is in contact with the optical contrast layer and provide a cavity distance/cavity size ratio of 3.125 or less, and (b) positioned opposite to the plurality of distinct antigens; 
 wherein the number and/or size of the cavities is selected to allow substantially complete hybridization of the antibody to at least one of the plurality of antigens within less than 60 minutes upon contacting the antigens with the antibody when oscillated at a frequency of 3.6 kHz; and 
 wherein a ratio between a number of the antigen spots to a number of cavities is at least 3:1, and wherein a ratio between an area of an antigen spot and cavity diameter is at least 1:3. 
 
     
     
       2. The microfluidic device of  claim 1  wherein the plurality of distinct antigens are bound to the optical contrast layer as a crude expression reaction. 
     
     
       3. The microfluidic device of  claim 2  wherein the crude expression reaction further comprises a detergent. 
     
     
       4. The microfluidic device of  claim 1  wherein the antigen spots comprise a dye selected from the group consisting of 3-amino-9-ethylcarbazole, 5-bromo-4-chloro-3-indolylphosphate, 3-3′-diaminobenzidine tetrachloride, 3,3′,5,5′-tetramethylbenzidine, and a colloidal metal. 
     
     
       5. The microfluidic device of  claim 1  wherein the carrier material and the optical contrast layer are configured to allow optical detection from opposite sides of the optical contrast layer. 
     
     
       6. The microfluidic device of  claim 1  wherein at least two of the distinct antigens have known quantified and known relative reactivities with respect to sera of a population infected with a pathogen expressing the antigens. 
     
     
       7. The microfluidic device of  claim 6  wherein the at least two of the distinct antigens are from the same pathogen. 
     
     
       8. The microfluidic device of  claim 1 , wherein a transparent topmost layer comprises the cavity layer, the device further comprising an imaging detector including a lens and configured to detect a secondary reagent bound to the antibody and the antibody hybridized to one of the plurality of distinct antigens bound to the optical contrast layer through the topmost transparent layer, wherein the optical contrast layer comprises nitrocellulose and is positioned at a bottom of the chamber. 
     
     
       9. The microfluidic device of  claim 1 , wherein cavities of the plurality of cavities have a diameter of about 100 μm. 
     
     
       10. The microfluidic device of  claim 1 , wherein the plurality of cavities comprises more than 1,000 cavities. 
     
     
       11. The microfluidic device of  claim 1 , wherein the porous optical contrast layer comprises a porous membrane, and wherein the porous membrane comprises a plurality of pores having sizes ranging from 0.2 μm and 0.5 μm.